Project Summary/Abstract Efforts to achieve a cure for HIV continue to be stymied by the persistence of inducible proviruses in latently infected cells. If the cells constituting the proviral reservoir can be induced to actively produce virus, then the reservoir could potentially be cleared by means of native immune killing or extrinsic therapies. Efforts in this direction to date have been hampered by an inability to reliably stimulate the entire latent reservoir such that this clearance can be achieved. One potential mechanism that could contribute to suboptimal latent cell activation and clearance is variation in the HIV Rev/RRE regulatory axis. Recent data from our laboratory have indicated that Rev/RRE functional activity varies greatly among primary isolates and that rather than acting as an on-off switch, Rev/RRE activity modulates viral replication. Upon re-activation, proviruses in latently infected cells with less functionally active Rev/RRE pairs would be expected to have decreased production of many viral proteins and virus particles, while continuing to produce normal levels of the immune modulatory viral Nef protein. Cells containing proviruses with low Rev/RRE activity may therefore be more difficult to clear in the so-called ?kick-and-kill? strategies for cure. Experiments in this proposal are designed to test this hypothesis. Subaims1a and 1b will utilize an established latency model and a set of engineered viruses that have altered Rev/RRE activity to directly assess how reactivation of latently infected cells with various latency reactivating agents affects viral activation and outgrowth. Subaim1c will measure Rev/RRE function in replication competent viruses isolated from patients' latent cells. These experiments will determine how Rev/RRE function varies compared to sequences derived from the peripheral blood of newly diagnosed patients, and whether there has been any selection with respect to levels of Rev/RRE functional activity. Specific Aim 2 will measure CTL killing of latency-reactivated cells to directly determine how Rev/RRE activity variation influences the susceptibility of the cells to immune elimination. Improved understanding of how the Rev-RRE regulatory system functions in viral latency may be one key to understanding why ?kick-and-kill? cure strategies have been unsuccessful to date. This information could in turn be used to optimize the ?kick? by modulating the Rev-RRE activity of latent virus. The proposed study thus has direct bearing on the most promising strategy to date to cure HIV and could provide crucial knowledge to advance this thus-far elusive goal.